In one aspect, the present invention relates generally to the field of ophthalmology and, more particularly, to compositions and methods for the prevention or treatment of eye diseases, such as cataracts. In another aspect, the present invention relates to the prevention or treatment of diseases caused by oxidative stress or having oxidative stress as a component.
The lens consists of concentric layers of fiber cells with hexagonal cross sections packed together to create a very regular array of fiber cells which stretch from anterior to posterior pole. The lens fiber cells lose all intracellular organelles that could contribute to light scattering during the process of differentiation and the ctyoplasmic protein concentration increases markedly. Lens transparency is the result of a uniform structure of the cells"" cytoplasm, which exists in an ordered, homogeneous state.
Approximately 35% to 60% of the total mass of the lens consists of structural proteins with the remainder being water. These cells contain primarily the crystallin proteins. When these proteins are modified by oxidative stress, conformational changes and aggregates result which, in turn, disrupt the protein lattice and damage the cell. This cellular damage leads to a further disruption of the regular layers of cells, resulting in opacities, or cataracts, of the lens.
More than 90% of the total lens protein consists of alpha, beta, and gamma crystallins (in excess of 300 mg/ml) in the lens cell cytoplasm. Crystallins are proteins containing numerous sulfhydryl groups, making them susceptible to oxidative damage. Protein S-thiolation, the formation of mixed disulfides between non-protein thiols and protein sulfhydryl groups in the lens, is an early event under oxidative damage (Lou et al., xe2x80x9cThe role of protein thiol mixed disulfides in cataractogenesis,xe2x80x9d Exp. Eye Res., vol. 50, 819-26 (1990)). Protein-glutathione (PSSG) and protein-cysteine (PSSC) are the products of such protein modification that have been implicated as a direct precursor of protein-protein disulfides (PSSP) in rat lenses cultured with H2O2. These changes ultimately lead to protein insolubility, loss of transparency of the lens tissue, and cataract formation (Cui et al., xe2x80x9cThe effect and recovery of long term H2O2 exposure on lens morphology and biochemistry,xe2x80x9d Exp. Eye Res., vol. 57, 157-67 (1993)).
In unstressed normal cells, the concentration of S-thiolated proteins is very low and the dethiolation rate, i.e., the breakdown of mixed disulfides by reduction of disulfide bonds, may be sufficient to maintain the fully reduced protein status. Such a situation has been observed in the H2O2 pre-exposed lenses after the oxidant was removed from the culture medium (Cui et al., Exp. Eye Res., vol. 57, 157-67 (1993); Lou et al., xe2x80x9cFurther studies on the dynamic changes of glutathione and protein-thiol mixed disulfides in H2O2 induced cataract in rat lenses: distributions and effect of aging,xe2x80x9d Curr. Eye Res., vol. 14, 951-58 (1995)). This implies that the maintenance of the redox status of the sulfhydryl groups of the protein is vital to the physiological function of the lens, a tissue abundant in sulfhydryl groups and extremely vulnerable to oxidative damage.
The redox state of the cell is maintained by many reductants, such as glutathione (GSH), NADPH, ascorbate, and tocopherol (Spector, xe2x80x9cThe lens and oxidative stress,xe2x80x9d in Oxidative Stress, Oxidants, and Antioxidants (Sies, Ed.) pp. 529-58, Academic Press: London (1991)). Among these reductants, GSH has a significant role in maintaining the reduced condition of the cell. Traditionally, GSH with glutathione reductase has been viewed as the system responsible for maintaining and regenerating the protein thiol groups that are susceptible to oxidation. Reduced GSH and other thiol containing molecules act, at least indirectly, as oxygen free radical scavengers, due to their easily oxidizable sulfhydryl groups and hence act as sulfhydryl antioxidants in normal lenses. Glutathione reductase regenerates GSH from its disulfide oxidized form (GSSG) and acts as a sulfhydryl buffering system, continually controlling the sulfhydryl/disulfide balance in the cell. GSH exists in great excess of its GSSG oxidized counterpart in the cellular cytoplasm, thus creating a strong potential for cytosolic proteins to exist in a sulfhydryl-reduced, non-crosslinked state. Deficiencies of GSH have been observed in cataractous lenses (Biochim et Biophys Acta, vol. 1138, 11-19 (1992)). This deficiency leads to a reduced ability of the cell to repair damage associated with oxidative stress.
Antioxidant therapy has been proposed to ameliorate the destructive effects of oxidation resulting in the formation of cataracts (American J. of Clinical Nutrition, vol. 53, pp.335S-345S (1991) and pp.352S-355S (1991)). Such proposed therapy has included the systemic administration of vitamins C and E and beta carotene. The use of phenolic antioxidants, such as probucol, to inhibit the development of cataracts, is disclosed in U.S. Pat. No. 5,061,734 to Mao et al. The use of N,Nxe2x80x2-bis(mercaptoacetyl)hydrazine derivatives as anticataract agents is disclosed in U.S. Pat. Nos. 5,686,450 and 5,688,828, both to Hellberg et al. PCT Application No. WO 94/03167 discloses the use of N-(3-mercapto-2,2-dimethylpropanoyl)cysteine and the intramolecular disulfide thereof for the treatment of cataract. Alpha-lipoic acid is disclosed as an anticataract agent in Maitra et al., xe2x80x9cAlpha-Lipoic Acid Prevents Buthionine Sulfoximine-Induced Cataract Formation in Newborn Rats,xe2x80x9d Free Radical Biology and Medicine, vol. 18, pp. 823-829 (1995). The use of phase separation inhibitors is proposed in U.S. Pat. No. 5,401,880 to Clark et al.
Cataracts are the leading cause of blindness in humans, with more than one million cataract extractions performed each year in the United States and with an estimated 5 to 10 million people becoming visually disabled each year due to cataracts. Cataracts in animals also pose a significant veterinary problem. Currently, no accepted nonsurgical therapy for the prevention and treatment of cataracts exists. Although various drugs have been proposed for use in the treatment and prevention of cataracts, it would be desirable to avoid potential adverse effects, such as chemical related toxicity, of such compounds.
The present invention is directed to compositions and methods for preventing or delaying the onset of cataracts and for treating or inhibiting the progression of cataracts in animals through the administration of thioltransferase (glutaredoxin) or derivatives thereof. In another aspect, the present invention is directed to the prevention and treatment of oxidative stress or damage and disease states associated therewith through the administration of thioltransferase or derivatives thereof. In one embodiment the animal may be a mammal. When the animal is a mammal, it may be a human.
The present invention is based on our discovery of the presence of thioltransferase in the lens and other ocular tissues, including the cornea cells, iris, and retina, all vulnerable to oxidative damage. Thioltransferase has been found by us to be an intrinsic repair system capable of dethiolating mixed disulfides associated with cataract initiation and formation, and thus is capable of repairing damaged proteins and nonprotein thiols by restoring them to their respective normally reduced states. It has also been found by us that thioltransferase can dethiolate enzymes damaged by thiolation, thus reactivating the catalytic function of the enzymes. Because thioltransferase in an endogenous biomolecule, it will cause no adverse effects due to chemical related toxicity.